Alliance Protein Laboratories, San Diego, CA, USA.
Eur Biophys J. 2023 Jul;52(4-5):233-266. doi: 10.1007/s00249-023-01629-0. Epub 2023 Feb 15.
Proper interpretation of analytical ultracentrifugation (AUC) data for purified proteins requires ancillary information and calculations to account for factors such as buoyancy, buffer viscosity, hydration, and temperature. The utility program SEDNTERP has been widely used by the AUC community for this purpose since its introduction in the mid-1990s. Recent extensions to this program (1) allow it to incorporate data from diffusion as well as AUC experiments; and (2) allow it to calculate the refractive index of buffer solutions (based on the solute composition of the buffer), as well as the specific refractive increment (dn/dc) of proteins based on their composition. These two extensions should be quite useful to the light scattering community as well as helpful for AUC users. The latest version also adds new terms to the partial specific volume calculations which should improve the accuracy, particularly for smaller proteins and peptides, and can calculate the viscosity of buffers containing heavy isotopes of water. It also uses newer, more accurate equations for the density of water and for the hydrodynamic properties of rods and disks. This article will summarize and review all the equations used in the current program version and the scientific background behind them. It will tabulate the values used to calculate the partial specific volume and dn/dc, as well as the polynomial coefficients used in calculating the buffer density and viscosity (most of which have not been previously published), as well as the new ones used in calculating the buffer refractive index.
要正确解释纯蛋白的分析超速离心(AUC)数据,需要辅助信息和计算来考虑浮力、缓冲液粘度、水合和温度等因素。自 20 世纪 90 年代中期推出以来,SEDNTERP 实用程序已被 AUC 界广泛用于此目的。该程序的最近扩展(1)允许它结合扩散和 AUC 实验数据;(2)允许它计算缓冲液的折射率(基于缓冲液的溶质组成),以及基于其组成的蛋白质的比折射增量(dn/dc)。这两个扩展对于光散射界以及 AUC 用户应该是非常有用的。最新版本还在部分比容计算中添加了新项,这应该会提高准确性,特别是对于较小的蛋白质和肽,并且可以计算含有重水同位素的缓冲液的粘度。它还使用了更新、更准确的水密度方程和棒和盘的流体动力特性方程。本文将总结和回顾当前程序版本中使用的所有方程及其背后的科学背景。它将列出用于计算部分比容和 dn/dc 的值,以及用于计算缓冲液密度和粘度的多项式系数(其中大部分以前未发表过),以及用于计算缓冲液折射率的新系数。